In general, acyl-CoA synthetase is an enzyme which thioesterifies the non-esterified fatty acid in the presence of CoA, ATP and Mg ion to make acyl-CoA, and it is also called thiokinase. This enzymatic reaction proceeds as follows: ##EQU1##
In recent years, with the solution of the physiological significance of non-esterified fatty acid in vivo, an increase and decrease of the amount of serum non-esterified fatty acid has come to be medically regarded as important. For example it has been found that an extreme increase of serum non-esterified fatty acid is recognized in case of disease of diabetes.
As a result, an increase of serum non-esterified fatty acid, detected by its determination, has come to be utilized for the diagnosis of the condition of such a disease as diabetes, but the serum non-esterified fatty acid has been generally determined by chemical colorimetric method. But the chemical colorimetric method requires a large quantity of blood, complicated analytical procedures and a longer time to perform it, resulting in this method being undesirable. Then, with the recent development of the method for clinical laboratory use, the quantitative determination method of non-esterified fatty acid by so-called enzymatic method has recently come to be used.
In order to quantitatively determine non-esterified fatty acid according to the enzymatic method, non-esterified fatty acid and acyl-CoA synthetase are reacted to form acyl-CoA and the product formed by this reaction is enzymatically determined thereby obtaining the concentration of non-esterified fatty acid. But with respect to the serum non-esterified fatty acid, since C.sub.16 -C.sub.18 long chain fatty acids are contained in large quantities in serum, an acyl-CoA synthetase that is able to thioesterify efficiently C.sub.16 -C.sub.18 long chain fatty acids to make acyl-CoA is naturally required. However, the acyl-CoA synthetase derived from liver microsome of rat is the only one which has been known which is fit for this purpose.
However, as this acyl-CoA synthetase is derived from an animal, it is very expensive; therefore, for economic reasons, it has been desired to derive the acyl-CoA synthetase from microorganisms which is not an expensive origin.
Heretofore, as the microorganisms being able to produce acyl-CoA synthetase, the following is known: Escherichia coli [European Journal of Biochemistry vol. 12, 576-582 (1970)]; Bacillus megaterium strain M [Biochemistry vol. 4, 85-95 (1965)]; Torulopsis Y.sub.8 [Journal of Bacteriology vol. 104, 1397-1398 (1970)]; Pseudomonas 22 [Journal of Bacteriology vol. 105, 1216-1218 (1971)] and Nocardia asteroides [Journal of Bacteriology vol. 114, 249-256 (1973)].
But since all of these known strains produce, as substrate specificity, such an acyl-CoA synthetase that has an optimum activity to the fatty acids having fourteen and less carbon atoms in the carbon chain, such enzyme derived from these strains can not be employed for clinical laboratory determination of C.sub.16 -C.sub.18 long chain fatty acids.